The body's immune system plays an important role in defending against infections caused by microorganisms such as bacteria, yeast, molds and viruses, against tumors and in the onset of allergies. However, the immune system is known to decrease in function due to aging, stress and illnesses such as cancer. Thus, there is a need for a safe and inexpensive immunoregulator that is highly effective for preventing microbial infections, demonstrating antitumor activity and preventing allergies.
Lactic acid bacteria are known to be safe microorganisms that are commonly consumed in the diet. In addition, they have also been reported to have various functionalities, such as intestinal regulatory action, serum cholesterol lowering action, immunoactivating action and immunoregulatory action such as antiallergic action (see, for example, Non-Patent Document 1). Examples of lactic acid bacteria commercially available in the form of probiotic lactic acid bacteria having immunoregulatory action include lactic acid bacteria belonging to the genera Streptococcus and Lactobacillus. These lactic acid bacteria are used to ferment dairy products (such as yogurt and yogurt drinks).
Immunoactivators and immunoregulators such as antiallergics having lactic acid bacteria as an active ingredient thereof are known in the prior art (see, for example, Patent Documents 1 to 12). Components reported to be involved with these immunoregulators having lactic acid bacteria as an active ingredient thereof include cell wall components in the form of peptidoglycans (see, for example, Non-Patent Document 2), lipoteichoic acid (see, for example, Non-Patent Document 3), lipoproteins (see, for example, Non-Patent Document 4), nucleic acids (see, for example, Non-Patent Documents 5 and 6) and heat shock proteins (see, for example, Non-Patent Document 7).
Among these related components, those reported to be nucleic acid components include the CpG motif (see, for example, Non-Patent Document 5) and the AT motif (see, for example, Non-Patent Document 6) of DNA.
At present, ten types of Toll-like receptors (TLR) are known to exist in humans. TLR are a type of protein present in the cell membrane that recognize extracellular pathogens and the like, and produce interferon and cytokines by transmitting that information inside cells.
In particular, TLR3 is known to recognize viral double-stranded RNA and induce interferon β promoter activation and interferon β production independent of MyD88. Interferon β activates dendritic cells and causes them to produce inflammatory cytokines such as interleukin 12 and TNF. Moreover, interleukin 12 establishes cellular immunity by inducing differentiation of naive T cells into type I helper T cells (Th1).
Double-stranded RNA refers to a structure uniquely observed in RNA viruses. Double-stranded RNA is formed when a virus infects host cells and replicates its virus genome by using host systems. In addition, double-stranded RNA is also present in the genome of double-stranded RNA viruses.
In addition, although reports on the subject are extremely rare, bacteria are also known to form double-stranded RNA under certain stressful conditions. In Escherichia coli, for example, a low molecular weight RNA known as RyhB is synthesized when the organism is subjected to iron ion depletion. This low molecular weight RNA is known to form partial base pairs with mRNA encoding iron-binding protein containing sodB mRNA, resulting in the formation of double-stranded RNA (see, for example, Non-Patent Document 8). However, whether or not this double-stranded RNA has an immunoregulatory action is unknown. In addition, there have thus far been no reports indicating the presence of double-stranded RNA in lactic acid bacteria.
As has been described above, although viral double-stranded RNA is considered to be preferable as an immunoactivator since it induces cellular immunity mediated by TLR3, use of the virus itself as an immunoactivator is not realistic in terms of safety, and modifications are required to ensure safety.
One solution involves the use of artificial double-stranded RNA for which safety has been ensured. In actuality, an artificial double-stranded RNA in the form of PolyI:PolyC has long been studied for use as an anticancer agent or antiviral agent due to its interferon-inducing effect. However, a system for enabling the required drug to be effectively incorporated into cells is required for general use in the body. For example, in order to allow PolyI:PolyC to act effectively in the body, it is necessary to incorporate it in the form of a liposome preparation (see, for example, Non-Patent Document 9).
In view of these circumstances, there is a need for a double-stranded RNA that can be used as an immuno activator, which is safe, and which is effectively incorporated into cells.
Patent Document 1: Japanese Unexamined Patent Application, First Publication No. H6-80575
Patent Document 2: Japanese Unexamined Patent Application, First Publication No. H9-227392
Patent Document 3: Japanese Unexamined Patent Application, First Publication No. H7-228536
Patent Document 4: Japanese Unexamined Patent Application, First Publication No. H10-167972
Patent Document 5: Japanese Unexamined Patent Application, First Publication No. H8-99887
Patent Document 6: Japanese Unexamined Patent Application, First Publication No. H5-252900
Patent Document 7: Japanese Unexamined Patent Application, First Publication No. 2003-113114
Patent Document 8: Japanese Unexamined Patent Application, First Publication No. 2004-26729
Patent Document 9: Japanese Unexamined Patent Application, First Publication No. 2004-18469
Patent Document 10: Japanese Unexamined Patent Application, First Publication No. 2000-95697
Patent Document 11: Japanese Unexamined Patent Application, First Publication No. H10-309178
Patent Document 12: Japanese Unexamined Patent Application, First Publication No. H9-2959
Non-Patent Document 1: Lactic Acid Bacteria Science and Technology, Japan Scientific Societies Press (1996)
Non-Patent Document 2: Lawrence C. and Nauciel C.: Infection and Immunity, 1998, 66, 4947-4949
Non-Patent Document 3: Cleceland M. G., et al.: Infection and Immunity, 1996, 64, 1906-1912
Non-Patent Document 4: Brightbill H. D., et al.: Science, 1999, 30, 732-736
Non-Patent Document 5: Krieg A. M., et al.: Nature, 1995, 374, 546-549
Non-Patent Document 6: Kitazawa H., et al.: Int. J. Food Microbiol., 2001, 65, 149-162
Non-Patent Document 7: Skeen M. J., et al.: Journal of Immunology, 1996, 156, 1196-1206
Non-Patent Document 8: Masse E. and Gottesman S., Proc. Natl. Acad. Sci. USA, 2002, 99, 4620-4625
Non-Patent Document 9: Nikkei Biotech, 552, 5 “Nippon Shinyaku Co. Ltd. Increased Dosage of the Double-Stranded Nucleic Acid Drug PolyI:PolyC Currently Being Tested in US”